Thermodynamic basis of site-specific cooperativity

Cooperative phenomena in biological macromolecules arise from the interaction of many distinct subsystems, such as structural domains or binding sites. Cooperative properties of the system as a whole, like protein folding or allosteric transitions, are subject to the restrictions imposed by thermodynamic stability. These restrictions, however, do not apply in the case of individual subsystems open to interactions with the rest of the macromolecule. The site-specific properties of such subsystems can be understood in general thermodynamic terms from those of a multicomponent system under particular conditions. The analogy provides a thermodynamic basis for site-specific Cooperativity. © 1994 John Wiley & Sons, Inc.     

Morphometric stepwise discriminant analysis of three genetically identified species within Pseudoterranova decipiens (Krabbe, 1878) (Nematoda: Ascaridida)

Univariate and multivariate statistical analyses have been performed on 19 morphometric variables of adult male specimens belonging to three genetically identified species within Pseudoterranova decipiens (Nematoda: Ascaridida) parasitic in the digestive tract of seals. Two morphometric keys are proposed for the identification of the three species. One key, which uses two variables, determines a frequency of error of 3.8% (3/79). The second key, which uses two canonical discriminant functions based on seven variables previously selected with a stepwise procedure, gives 100% (76/76) accurate classification.     

Interaction of trace elements in a longitudinal study of human milk from full-term and preterm mothers

Concentrations of 8 trace elements (Fe, Cu, Zn, Se, Br, Pb, Rb, and Sr) at different lactation time were measured by the PIXE multi-elemental technique. Time dependence and interelement correlations were studied. A total of 200 milk samples from 32 lactating mothers were supplied from 2 to 120 d after delivery of 26 full-term and 6 preterm infants. All elements showed a lognormal frequency-distribution. The Fe, Cu, Zn, and Se contents in preterm milk were found to be somewhat different with respect to full-term milk. Cu, Zn, Se, Br, Pb, and Rb concentrations declined with lactation time, both in pre- and full-term samples. Sr and Fe contents did not show any change with time. Detailed analysis of data by partial correlation and multiple regression methods was performed. No substantial differences between preterm and full-term samples were found in the results of partial correlation analysis. Cu and Zn were found to be correlated with lactation time, whereas the measured time dependence for the other elements has to be attributed to the effect of the existing interelement correlation. All the measured elements appeared to be correlated with at least one other element. In particular, Se was inversely correlated with Zn and directly with Cu. The zinc and copper contents in milk can therefore depend on the variation in the mother selenium intake.     

Toxins of Pseudomonas syringae pv. syringae affect H+-transport across the plasma membrane of maize

The activity of some phytotoxic metabolites of Pseudomonas syringae pv. syringae Van Hall strains B359 and B301 on in vivo and in vitro systems of H⁺-transport across the plasma membrane of maize (Zea mays L., hybrid Paolo) was investigated. In particular syringomycin, the first lipodepsinonapeptide isolated from Pss and already studied in plants and yeasts for its effects on several physiological systems, was compared with the recently described lipodepsipeptides with 22 or 25 amino acid residues, so called syringopeptins. The in vivo activity of the phytotoxins was tested on fusicoccin-stimulated H^?-extrusion from cuttings of maize roots, which was inhibited by both types of toxins, with syringomycin more efficient than the syringopeptins. In vitro the H⁺-ATPase activity of predominantly right-side-out plasma membrane vesicles purified by two-phase partitioning was stimulated by 10 μM syringomycin and inhibited by higher levels, in agreement with the results of others with preparations of dicotyledons. Also the inhibition of the phosphohydrolytic activity of inside-out vesicles of mung bean plasma membrane was confirmed for maize. In both types of vesicles the syringopeptirts were better inhibitors than syringomycin. The pH gradient formed on addition of ATP to predominantly (25% latency) inside-out vesicles was immediately and completely collapsed by syringomycin and syringopeptins; H⁺-pumping was prevented if the toxins were added before ATP. The inhibition was concentration dependent, but at very low concentrations the effect was inverted. The results of the present investigation, carried out with maize preparations, confirm and extend the evidence so far obtained with dicotyledons in favour of the plasma membrane as an important site of interaction of syringomycin with the plant cell. They also indicate that, except for some details, the effects of syringopeptins at the level of the plasma membrane are the same as those of syringomycin.     

Large DNA inversions,deletions, and TaqI site mutations in severe haemophilia A

Large DNA inversions caused by an intrachromosomal recombination between homologous regions located in intron 22 and 5 of the factor VIII gene have recently been identified in patients with severe haemophilia A. To evaluate better the prevalence of this large inversion and to estimate the overall sensitivity of the Southern blot/hybridization method we analysed the factor VIII gene of 49 unrelated patients with severe haemophilia A. All patients were screened for the inversion mutation, TaqI site mutations, and deletions. Mutations were identified in 31 (63%) patients, and comprised 24 large inversions, 4 partial deletions, and 3 point mutations. Three different haplotypes were characterised in the patients presenting the inversion mutation, confirming its independent origin. Two novel deletions are reported: a large one spanning from intron 14 to intron 22 and a deletion of 86 bp comprising the 3 region of exon 1 and 39–41 bp of intron 1. DNA sequencing of the deletion junction showed no significant homology between normal 5 and 3 sequences around the breakpoints. A novel missense mutation is also reported: CGAGGA, Arg-2209 to Gly. These results confirm that the inversion mutation is the most common cause of severe haemophilia A and indicate that the Southern blot/hybridization assay should be used as the first method for screening of mutations in severe haemophilia A.     

H1° and H3.3B mRNA levels in developing rat brain

Two overlapping rat cDNAs, covering a continuous region of 1107 base pairs, have been isolated and sequenced. The clones contain identical open reading frames, encoding a 136 amino acid long polypeptide which exhibits 100% identity to other mammalian H3.3 histone variants. We show that the inserts derive, in particular, from the H3.3B gene. We used these inserts and an insert from an H1° encoding clone, previously described (6), as probes to study the accumulation of mRNAs encoding the corresponding histone replacement variants (namely, H1° and H3.3) during rat brain development. We found that the concentration of both H1° and H3.3B mRNAs decreases from the embryonal day 18 (E18) to the postnatal day 10 (P10), with inverse correlation to protein accumulation.This paper is dedicated to our friend Paolo Carbone who devoted his life to research and teaching in Genetics. We will always remember him for scientific honesty and for his unique qualities of humanity.     

Coccoid Helicobacter pylori Not Culturable In Vitro Reverts in Mice

An experimental rodent model was used to demonstrate the viability of the coccoid form of Helicobacter pylori. Concentrated suspensions were prepared for the two different morphologies: at 2 days incubation for the bacillary forms and at 20 days incubation for the “dormant” forms. The strains used for incubation were two fresh isolates from humans with duodenal ulceration, and two collection strains. Five hundred microliters of culture (OD₅₅₀ = 5 Mc Farland) of Helicobacter pylori with bacillary (2-5×10⁹ CFU/ml) and coccoid (0 CFU/ml) morphology were inoculated intragastrically in BALB/c mice. The gastric mucosa of the mice was colonized by Helicobacter pylori with the administration of fresh bacillary and coccoid cultures and not with the established cultures. Helicobacter pylori was isolated at 1 week after inoculation with the administration of fresh bacillary cultures, while fresh coccoid Helicobacter pylori was recovered in mice stomachs after 2 weeks of inoculation. After colonization, histopathologic changes occurred after 1 month from inoculation; all colonized mice showed a systemic antibody response to Helicobacter pylori. These results support the thesis of the viability of coccoid Helicobacter pylori non-culturable in vitro and confirm that concentrated bacterial suspensions are able to colonize and to produce gastric alterations in this suitable animal model.